Proton translocation coupled to trimethylamine N-oxide reduction in anaerobically grown Escherichia coli

Abstract

Proton translocation coupled to trimethylamine N-oxide reduction was studied in E. coli grown anaerobically in the presence of trimethylamine N-oxide. Rapid acidification of the medium was observed when trimethylamine N-oxide was added to anaerobic cell suspensions of E. coli K-10. Acidification was sensitive to the proton conductor 3,5-di-tert-butyl-4-hydroxybenzylidenemalononitrile (SF6847). No pH change was shown in a strain deficient in trimethylamine N-oxide reductase activity. The apparent H+/trimethylamine N-oxide ratio in cells oxidizing endogenous substrates was 3-4 g-ions of H+ translocated/mol trimethylamine N-oxide added. The addition of trimethylamine N-oxide and formate to EDTA-treated cell suspension caused fluorescence quenching of 3,3''-dipropylthiacarbocyanine [diS-C3(5)], indicating the generation of membrane potential. The reduction of trimethylamine N-oxide in E. coli is apparently catalyzed by an anaerobic electron transfer system, resulting in formation of a proton motive force. Trimethylamine N-oxide reductase activity and proton extrusion were also examined in chlorate-resistant mutants. Reduction of trimethylamine N-oxide occurred in chlC, chlG and chlE mutants, whereas chlA, chlB and chlD mutants, which are deficient in the Mo cofactor, could not reduce it. Protons were extruded in chlC and chlG mutants, but not in chlA, chlB and chlD mutants. Trimethylamine N-oxide reductase activity in a chlD mutant was restored to the wild-type level by the addition of 100 .mu.M molybdate to the growth medium, indicating that the same Mo cofactor as used by nitrate reductase is required for the trimethylamine N-oxide reductase system.